Bl21 codonplus ril cells

Manufactured by Agilent Technologies

BL21-CodonPlus-RIL cells are competent cells designed for high-level protein expression. They are optimized for expression of proteins that contain rare codons.

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Lab products found in correlation

Bl21 star de3 cells, by Thermo Fisher Scientific (1 mentions) Dnase 1, by Merck Group (1 mentions) Lysozyme, by Merck Group (1 mentions) Histrap column, by GE Healthcare (1 mentions) Mbptrap hp column, by GE Healthcare (1 mentions) E coli bl21 star de3 cells, by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

BL21-CodonPlus(DE3)-RIL cells (27 citations) BL21-CodonPlus (DE3)-RIL competent cells (21 citations) BL21-CodonPlus-RIL (9 citations) BL21-CodonPlus cells (8 citations) BL21-CodonPlus (DE3)-RIL E. coli cells (6 citations) Escherichia coli BL21-CodonPlus (DE3)-RIL cells (4 citations) BL21 (CodonPlus) RIL competent cells (2 citations)

5 protocols using bl21 codonplus ril cells

Purification and Histone Methyltransferase Assay of HAP

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Full length open reading frame of HAP was amplified from the pGWB402 vector using the primers listed in Supplementary Table S5 and cloned into the pGEX4T-3 expression vector. The resulting plasmid was transformed into E. coli BL21-CodonPlus-RIL cells (Agilent) and protein expression assays were then performed as previously described [58 (link)]. Briefly, cells containing pGEX4T-3 vector were grown to an exponential phase and protein expression was induced by adding IPTG. Bacteria were then disrupted by sonication and protein purification was performed using GST-sepharose (Pharmacia) according to the manufacturer’s instructions. Finally, Histone H3 (K4, K9 and K27) methyltransferase activity was determined using HMT activity quantification assay kits (Abcam) following the manufacturer’s instructions.

Fonseca R., Capel C., Yuste-Lisbona F.J., Quispe J.L., Gómez-Martín C., Lebrón R., Hackenberg M., Oliver J.L., Angosto T., Lozano R, & Capel J. (2022). Functional characterization of the tomato HAIRPLUS gene reveals the implication of the epigenome in the control of glandular trichome formation. Horticulture Research, 9, uhab015.

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HIV-1 Capsid Protein Purification

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E.coli BL21 CodonPlus-RIL cells (Agilent Inc. Santa Clara, CA) transformed with pGEX-4T-3 or pGEX-HIV-CA were used for purification of GST proteins using standard methods. Fifteen independent synthetic peptides covering HIV-1 CA were designed and provided by Sigma-Aldrich (Sigma-Aldrich Co, St. Louis, MO). The amount of CA protein was quantified using HIV-1 CA (p24) enzyme-linked immunosorbent assay kits (ZeptMetrix Corporation, Buffalo, NY).

Takeuchi H., Saito H., Noda T., Miyamoto T., Yoshinaga T., Terahara K., Ishii H., Tsunetsugu-Yokota Y, & Yamaoka S. (2017). Phosphorylation of the HIV-1 capsid by MELK triggers uncoating to promote viral cDNA synthesis. PLoS Pathogens, 13(7), e1006441.

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Purification of Pf LipL1 Protein Variants

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All constructs were transformed into BL21-Star (DE3) cells (Invitrogen) containing the pRIL plasmid isolated from BL21-CodonPlus-RIL cells (Agilent) and plasmid pRK586 encoding the Tobacco Etch Virus (TEV) protease as described.^{17 (link)} These cells produce a protein product fused to an amino-terminal hexahistidine tag. 2L of TB media containing ampicillin, kanamycin, and chloramphenicol were inoculated with an overnight culture for an initial OD₆₀₀ of approximately 0.1. The cells were grown to mid log phase at 37°C and then the temperature was reduced to 20°C. Protein expression was induced with 0.4 mM Isopropyl β-D-1-thiogalactopyranoside (IPTG) and the cells were harvested after 10h. Purification was performed as described previously for full-length PfLipL1.¹⁴ Briefly, PfLipL1 variants were purified by immobilized metal ion chromatography followed by cation exchange chromatography and gel filtration chromatography. The first two steps of purification were performed on the same day to avoid proteolytic cleavage. Purified PfLipL1 mutants were concentrated to approximately 5 mg·mL⁻¹ and stored at −80°C.

Guerra A.J., Afanador G.A, & Prigge S.T. (2017). Crystal structure of lipoate-bound lipoate ligase 1, LipL1, from Plasmodium falciparum. Proteins, 85(9), 1777-1783.

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Purification of pMAL-cHT PyrKII Protein

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The pMAL-cHT PyrKII protein was purified from BL21 Star (DE3) E. coli cells (ThermoFisher) also harboring the pRIL plasmid from BL21-CodonPlus-RIL cells (Agilent). Cells harboring both plasmids were grown in LB medium, and shaken at 220 rpm at 37°C until an OD₆₀₀ of 0.6 was reached. At this point, protein expression was induced with 0.4 mM isopropyl β-thiogalactoside (IPTG) and the cells were then incubated for an additional 4 hr at 27°C. Cells were lysed with 20 mM Tris-HCl at pH 7.5, 100 mM KCl, 1 mg/mL lysozyme (Sigma), 2.5 µg/mL DNAse I (Sigma), and 1 mM PMSF, followed by sonication. The MBP-PyrKII protein was purified from the resulting lysate using a 5 mL MBPTrap HP column (GE Healthcare), and eluted with 100 mM maltose. The resulting protein was then cleaved with TEV protease and further purified using a 5 mL HisTrap column (GE Healthcare). The protein was then eluted with an imidazole gradient, and peak fractions were collected, pooled, and concentrated for storage in 20 mM Tris-HCl (pH 7.5), 100 mM KCl, 10% glycerol at −80°C.

Swift R.P., Rajaram K., Keutcha C., Liu H.B., Kwan B., Dziedzic A., Jedlicka A.E, & Prigge S.T. (2020). The NTP generating activity of pyruvate kinase II is critical for apicoplast maintenance in Plasmodium falciparum. eLife, 9, e50807.

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Expression of Proteins in E. coli

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All proteins were expressed in E. coli BL21-CodonPlusRIL cells (Agilent) in Luria Broth in a 2 L fermenter (Sartorius Stedim) operated at 37°C.

Eren E., Watts N.R., Dearborn A.D., Palmer I.W., Kaufman J.D., Steven A.C, & Wingfield P.T. (2018). Structures of Hepatitis B Virus core- and e-antigen Immune-complexes Suggest Multi-Point Inhibition. Structure (London, England : 1993), 26(10), 1314-1326.e4.

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